• Korean Journal of Soil Science and Fertilizer
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• v.43 no.4
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• pp.490-497
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• 2010

A chitinolytic bacterium having a strong antagonistic activity against various pathogens including Phytophtora capsici was isolated from rhizosphere soil, and identified as Lysobacter enzymogenes (named as LE429) based on 16S rRNA gene sequence analysis. This strain produced a number of substances such as chitinase, ${\beta}-1$, 3-glucanase, lipase, protease, gelatinase and an antibiotic compound. This antibiotic compound was purified by diaion HP-20, sephadex LH-20 column chromatography and HPLC. The purified compound was identified as phenylacetic acid by gas chromatography-electron ionization (GC-EI) and gas chromatography-chemical ionization (GC-CI) mass spectrometry. In field experiment, pepper plants were treated by the strain LE429 culture (CB), neem oil solution (NO), combination (CB+NO) or control (CON). Plant height and number of branches, flowers and pods of pepper plant in CB treatment were generally highest, and followed by CB+NO, CON and NO. The fungal pathogens were strongly inhibited, while several insect pests were discovered in CB treatment. Any insect pests were not found, while all fungal pathogens tested were not suppressed in NO treatment. However, in CB+NO treatment, non incidence of fungal pathogens and insect pests were found. The strain LE429 producing secondary metabolites with neem oil should be a potential agent to control fungal diseases and insect pests.

• Korean Journal of Agricultural Science
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• v.14 no.2
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• pp.286-294
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• 1987

This experiment was carried out to investigate the bacteriolytic action of the egg white lysozyme isolated from Korean native Ogol fowl and to obtain the data for utilization of the enzyme as a food preservative. Staphyococcus aureus phage type 29 and Bacillus subtilis ATCC 6633 among the microorganisms tested were lyzed by the treatment with 0.05% lysozyme, but Staphylococcus aureus phage type 57 in addition to E. coli etc. was found to be a lysozyme- insensitive species. The lysis of S. aureus phage type 29 was maximized when incubated in nutrient broth (pH 7.0) at $37^{\circ}C$ for 24 hours and suspended it to absorbance 0.6 at 540nm in 0.05M sodium acetate but fer (pH 4.5) and then treated it with the 0.05% lysozyme for 30 min. at $30^{\circ}C$. It was found that the effect of 0.05% lysozyme in combination with 1% glycine on the growth inhibition of S. aurecus phage type 29 increased more 50% than that in the absence of glycine, but not effect with other any additeves and metal ions tested.

• The Korean Journal of Mycology
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• v.23 no.2 s.73
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• pp.129-138
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• 1995

DNA fragment being able to restore in vitro activity of ${\beta}-1,3-glucan$ synthase was cloned by transformation of the Saccharomyces cerevisiae LP353 mutant strain with genomic library constructed in the YCp50. For the selection of transformants which showed no detectable phenotype linked to recovery of the defect in ${\beta}-1,3-glucan$ synthase activity, the colony autoradiography was succesfully applied. The restriction map of the cloned DNA fragment, which is 8.5-kb in length, was constructed. Both the YEplac195 and the YCp50 carrying the 8.5-kb fragment increased ${\beta}-1,3-glucan$ synthase activity of LP353 by two fold. Neither the YEplac195 nor the YCp50 carrying the 8.5-kb DNA fragment, however, complemented the temperature-dependent osmotic sensitivity which is another distinctive phenotype of LP353. Subcloning experiments indicated that a functional region was located in 4.8-kb BglII-KpnI fragment. The 4.8-kb fragment was also able to increase the level of ${\beta}-1,3-glucan$ content in cell wall as well as the resistance of cells to cell wall lytic enzyme, ${\beta}-1,3-glucanase$. The growth rate of the LP353 with 4.8-kb fragment was almost same as that of wild type strain in liquid medium with 1.2 M sorbitol at nonpermissive temperature. Taken these results together, the 4.8-kb fragment seemed to contain the BGS2 gene for ${\beta}-1,3-glucan$ synthase activity in yeast S. cerevisiae.

• Applied Biological Chemistry
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• v.41 no.2
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• pp.125-129
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• 1998

Xanthomonas sp. isolated from soil exhibited cell wall lytic activity of Candida albicans and secreted chitinase in chitin media. Especially, the chitinase activity was induced by chitin and reached a maximum level at 3 days culture in chitin media. We constructed genomic library of Xanthomonas sp. using cosmid vector in E. coli. Oligonucleotide probe was synthesized from the consensus sequence corresponding to chitinase active site, which was derived from the comparison of amino acid sequences of bacterial chitinase genes. Using this oligonucleotide probe, we screened the genomic library. By restriction enzyme mapping of the positive clones, we identified 4 independent clones which may contain the chitinase gene. One of the clones, named pXCH1 (1.2 kb insert), was further analyzed. Northern blot analysis indicated that is transcripts, 1 kb and 0.8 kb, were induced by chitin. When the cloned gene was induced by IPTG in E.coli cell, chitinase activity which was secreted onto culture media was not observed. However, when the cell was disrupted by using sonicator and then centrifuged, the supernatant exhibited chitinase activity. SDS-PAGE of the supernatant indicated that about 35 kDa protein was induced by IPTG. From these results, it was concluded that the cloned DNA was one of the chitinase genes of Xanthomonas sp.

• Korean Journal of Agricultural Science
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• v.13 no.2
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• pp.185-192
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• 1986

Some factors affecting the protoplast release from mycelia of Ganoderma lucidum and regeneration of the protoplast were investigated and the results obtained are summarized as follows; Novozym 234 as a lytic enzyme was the most effective for the protoplast release from mycelia of Ganoderma lucidu m and its optimal concentration was 10mg per ml of osmotic stabilizer. The highest number of protoplasts were released after 3 hours incubation in the reciprocal shaking bath at 120 oscillations a minute. Among six osmotic stabilizers tested, 0.6M sucrose showed the best result. SCM medium showed good mycelial growth and high yields of protoplasts. The protoplasts released from the mycelium of G. lucidum were regenerated at 0.20 to 0.27 percent on MCM, MMM and SCM. Of the cultures obtained from protoplasts regenerated, 13 to 29 percent were monokaryon.

• Korean Journal of Plant Tissue Culture
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• v.25 no.2
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• pp.109-113
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• 1998

In this study we tried to transform an antimicrobial peptide gene (Shiva) under the promoter of tomato phenylalanine ammonia-lyase (tPAL5) into tobacco and potato plants. Antimicrobial peptide gene was isolated originally from giant silk moth (Hyalophora cecropia) and modified ie nucleotide sequence to increase antimicrobial activity. Transgenic tobacco plants were regenerated and their seeds were tested on the media containing kanamycin (500 mg/L). The results of PCR amplification and genomic Southern blot hybridization confirmed the integration of construct (tPAL5 promoter-Shiva-NOS-GUS-NOS) into chromosome. We observed that one of the transgenic tobacco plants showed chromosome rearrangement when integrated. In case of potato transformation, the efficiency of regeneration was maximized at the medium containing Zeatin 2mg/L, NAA 0.01mg/L, GA$_3$ 0.1mg/L. We also observed the high expression of GUS (${\beta}$-glucuronidase) enzyme which was located next to the terminator sequence of nopaline synthase gene (NOS) in the vascular tissue of stem, leaves of transgenic potatoes. This result suggested that a short sequence of Shiva gene (120 bp) and NOS terminator sequence might be served as a leader sequence of transcript when translated.

• Korean Journal of Food Science and Technology
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• v.12 no.4
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• pp.254-262
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• 1980

A series of experiment were carried out to separate the factor accelerating the lysis of cell wall of $Saccharomyces\;sak{\acute{e}}$ from the preparation of crude zymolyase obtained from Arthrobacter luteus. An attempt was also made to purify the enzyme which is essential for the study on the separation of the factor. The results are summarized as follows: 1. Crude zymolyase was fractionated 5 peaks $(A{\sim}E)$ containing three peaks $(A{\sim}C)$ passed through the column by the chromatography on Biogel CM-30. 2. Among the five peaks, peak E (protease fraction) was found to contain the factor accelerating the lytic activity of the zymolyase. 3. L-c fraction purified in almost free form from the nonlytic ${\beta}-1$, 3-glucanase, protease and inert protein by the affinity adsorption chromatography with Sephadex G-75 gel was obtained from zymolyase fraction (peak D). When it was subjected to polyacrylamide gel disc electrophoresis, only one clear protein band was observed at pH 4. 5, but still detected two or more band at pH 8. 3.

• Korean Journal of Soil Science and Fertilizer
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• v.44 no.6
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• pp.1150-1157
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• 2011

Streptomyces sampsonii KK1024 having strong chitinolytic activity was isolated from crab-shell rich soil at Muan, Jeolanamdo. The KK1024 produced chitinase, protease, gelatinase and lipase. When 50% of KK1024 culture broth was treated to juveniles and eggs of root-knot nematode, juvenile mortality at 3 days was 81.67% and egg hatch rate at 5 days was 2.00%. When $183.7{\mu}g\;mL^{-1}$ of crude enzyme produced by KK1024 was treated, juvenile mortality at 3 days was 96.00% and egg hatch rate at 5 days was 5.33%. At 1% of butanol extract from KK1024, juvenile mortality was highest with 90.00% and egg hatch rate was lowest with 0%. The comparison of the effect of KK1024 culture broth with only medium, synthetic fertilizer, and commercial nematicide on tomato growth and nematode infection was examined in pot trials. KK1024 culture broth showed lower number of egg mass and gall in plant, and population of juveniles in soil compared with only medium and synthetic fertilizer treatment, but not in commercial nematicide. However, the highest shoot weight and length was discovered in KK1024 culture broth. These results suggest that Streptomyces sampsonii KK1024 producing lytic enzymes and nematicidal compounds can be one of candidates for biocontrol agents against root-knot nematodes.